Discover the Blyscan™ Glycosaminoglycan Kit: Your go-to Solution for reliable sGAG and Proteoglycan Analysis! Our versatile, widely cited kit utilizes a dye-binding approach to quantitatively measure sulfated glycosaminoglycans (sGAG) and proteoglycans in cells, tissues, and fluids from in-vivo and in-vitro sources.
Description
The Biocolor Blyscan™ Glycosaminoglycan (sGAG) Assay enables quantitative measurement and analysis of sulfated proteoglycans and glycosaminoglycans (sGAG).
Soluble sGAGs may be assayed directly, whereas solid materials will require a preliminary papain extraction.
The assay can be used to measure the total sGAG content and can also be adopted to determine the O- and N-sulfated glycosaminoglycan ratio within test samples.
Suitable Samples
In-vivo:
- Solid samples, including cartilage, bone, connective tissue, tumor tissue
-
Liquid samples, including fluids such as urine, amniotic fluid, or synovial fluid
In-vitro:
- Solid samples, such as deposited ECM on 2D/3D culture surfaces by enzymatic treatment
- Liquid samples, Culture media during 2D/3D cell culture
The assay requires that sulfated polysaccharides or sGAGs are in a soluble form. A preliminary enzymatic extraction step is required for solid samples (enzyme not supplied with kit).
The assay is not suitable for use with samples containing alginates or that comprise degraded sulfated disaccharide fragments.
Principle of Method
Colormetric Assay, Dimethylmethylene Blue Assay (DMMB)
The dye label used in the assay is 1,9-dimethylmethylene blue (DMMB) and the dye is employed under conditions that provide a specific label for the sulfated polysaccharide component of proteoglycans or the protein free sulfated glycosaminoglycan chains.
Step 1. Blyscan dye reagent contains DMMB dye in an optimised buffer. Addition of Dye reagent to samples containing sGAG results in the formation of a dye/sGAG complex due to a charge interaction between dye and GAG sulfate groups.
Step 2. Over a 30 minute incubation Dye-labelled sGAGs precipitate out of solution and are collected by centrifugation. Following removal of unbound dye, the remaining bound dye is released from the complex by addition of dye dissociation reagent. Released dye is quantified spectrophotometrically.
Step 3. The sGAG content of unknown samples may be quantified by comparison against a calibration curve prepared using a standard of purified Chondroitin-4-sulfate supplied with the kit.
The Blyscan Dye reagent is formulated to minimize binding to other charged sample components such as nucleic acids, a problem with some older dye-based sGAG assays.
Test Sample Composition
An extensive range of mammalian proteoglycans have been measured using this assay. The following sGAG, either still attached to the peptide/protein core, or as free chains can be assayed:
- chondroitin sulfates (4- & 6-sulfated)
- keratan sulfates (alkali sensitive & resistant forms)
- dermatan sulfate
- heparan sulfates (including heparins)
Hyaluronate is not detected by the Blyscan Assay. The presence of hyaluronate in test samples does not interfere with the measurement of sGAG.
Specifications
|
Detection Method |
Colorimetric Detection (656 nm) |
| Assay Rangehttps://cdn.shopify.com/s/files/1/0563/4487/1064/files/Blyscan-manual-webopt.pdf?v=1672706553 | 0 - 50 µg/ml |
| Limit of Detection | 2.5 µg/ml |
| Sensitivity | 0.5 µg |
| Assay Run-Time |
1 hour |
| Measurements Per Kit | 110 in total (allows a maximum of 48 samples to be run in duplicate alongside a standard curve) |
Kit Contents
The standard size Blyscan™ Glycosaminoglycan kit contains:
- Blyscan Dye Reagent (1x 110 ml)
- sGAG Reference Standard (1x 5 ml, 100 µg/ml Bovine tracheal chondroitin 4-sulfate)
- Dissociation Reagent (1x 110 ml)
- Sodium Nitrite (1x 15 ml)
- Acetic Acid (1x 15 ml)
- Ammonium Sulfamate (1x 15 ml)
- 1.5 ml micro-centrifuge tubes for dye-labelling reaction
- Assay kit manual
NOTE: Additional reagents may be required for sample preparation prior to assay. Consult manual or contact us for further details.
Applications
- fibrous and hyaline cartilages
- arteries, heart, lung, skin and other material containing extracellular matrix (connective tissue), and solid tumors
- extracellular matrix components that may be released by live cells into culture medium
- soluble sGAG from synovial fluid, urine and gel chromatography fraction aliquots
Technical Video
Other Names/Synonyms
Blyscan sGAG Assay, s-glycosaminoglycan, sulfated glycosaminoglycan
Resources
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Ilex Life Sciences LLC is an official distributor of Biocolor Ltd.